中国组织工程研究

中国组织工程研究 ›› 2011, Vol. 15 ›› Issue (27): 4959-4963.doi: 10.3969/j.issn.1673-8225.2011.27.005

• 骨髓干细胞 bone marrow stem cells • 上一篇    下一篇

肝纤维化大鼠血清体外诱导骨髓间充质干细胞向肝细胞的分化

范莎莎，石詠中，袁友红，曹友德   

  1. 湖南省人民医院临床医学研究所肝细胞移植中心，湖南省长沙市410005
  • 收稿日期:2011-02-16 修回日期:2011-04-21 出版日期:2011-07-02 发布日期:2011-07-02
  • 通讯作者: 曹友德，主任技师，湖南省人民医院临床医学研究所肝细胞移植中心，湖南省长沙市410005
  • 作者简介:范莎莎★，女，1982年生，湖南省衡阳市人，汉族，2008年中南大学毕业，硕士，主要从事干细胞分化的研究。 fresh0225@126.com
  • 基金资助:

    湖南省科技厅课题(2010TD2031)。

In vitro differentiation of bone marrow mesenchymal stem cells induced by liver cirrhosis sera of rats into hepatocytes

Fan Sha-sha, Shi Yong-zhong, Yuan You-hong, Cao You-de   

  1. Liver Cell Transplantation Center, Institute of Clinical Medicine, Hunan Provincial People’s Hospital, Changsha  410005, Hunan Province, China
  • Received:2011-02-16 Revised:2011-04-21 Online:2011-07-02 Published:2011-07-02
  • Contact: Cao You-de, Chief technician, Liver Cell Transplantation Center, Institute of Clinical Medicine, Hunan Provincial People’s Hospital, Changsha 410005, Hunan Province, China fresh0225@sina.com
  • About author:Fan Sha-sha★, Studying for master’s degree, Liver Cell Transplantation Center, Institute of Clinical Medicine, Hunan Provincial People’s Hospital, Changsha 410005, Hunan Province, China fresh0225@126.com
  • Supported by:

    the Project of Hunan Science and Technology Bureau, No. 2010TD2031*

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摘要：

背景：近年来临床运用骨髓间充质干细胞移植治疗晚期肝纤维化取得了较好疗效，但由于肝源稀少，骨髓干细胞体外分化为成熟肝细胞的技术仍然不成熟。
目的：探讨骨髓间充质干细胞体外诱导其分化为肝细胞的可行性。
方法：构建大鼠肝纤维化模型，分离、纯化并鉴定大鼠骨髓间充质干细胞，制备正常及肝纤维化大鼠血清，取第3代骨髓间充质干细胞分组培养，分别加入以下培养基：DMEM+体积分数10%胎牛血清；肝细胞生长培养基(HGM)+体积分数5%胎牛血清；HGM+5%正常大鼠血清；HGM+5%肝纤维化大鼠血清；HGM+5%肝纤维化大鼠血清+25 μg肝细胞生长因子。观察各组细胞形态变化，MTT法测定细胞生长曲线，采用Realtime-PCR检测甲胎蛋白和细胞角蛋白18的表达，溴甲酚绿法检测上清液中白蛋白水平，ELISA法检测培养上清液中转化生长因子β1表达。
结果与结论：正常大鼠血清能促进骨髓间充质干细胞的增殖；肝纤维化大鼠血清对骨髓间充质干细胞促增殖作用最好，且能有效诱导其向肝细胞分化，联合使用肝细胞生长培养基能提高分化率。

关键词: 肝纤维化, 肝细胞生长因子, 骨髓间充质干细胞, 转化生长因子&beta, 1, 大鼠

Abstract:

BACKGROUND: In recent years, the clinical use of bone marrow mesenchymal stem cells (BMSCs) transplantation for the treatment of liver fibrosis has achieved good effects, but the source of the liver is rare. The techniques for bone marrow stem cells differentiating into mature hepatocytes are still immature.
OBJECTIVE: To discuss the possibility of the differentiation of BMSCs induced by liver cirrhosis sera of rats in vitro into hepatocytes．
METHODS: Rat BMSCs were isolated, purified and identified; normal and liver cirrhosis sera of rats were prepared; the third passage BMSCs were divided into groups for culture: Group A: DMEM+10% fetal bovine serum (FBS); Group B: hepatocyte growth medium (HGM)+5% cirrhosis sera of rats; Group C: HGM+5% normal rat sera; Group D: HGM+5% liver cirrhosis sera of rats; Group E: HGM+5% liver cirrhosis sera of rats+25 μg hepatocyte growth factors (HGF). Cell morphology was observed and the cell growth curve was measured with MTT assay. The expressions of Alpha-fetoprotein and Cytokeratin18  were detected by reverse transcription polymerase chain reaction, and the albumin content in supernatant was determined by bromoeresol green method. Transforming growth factors β1 in supernatant was detected by ELISA.
RESULTS AND CONCLUSION: Normal rat sera can obviously promote the proliferation of BMSCs; liver cirrhosis sera of rats are of certain effect on BMSCs proliferation and can effectively induce the differentiation of BMSCs into hepatocytes. The combined application with HGF can raise BMSCs differentiation.

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